Formulation for regeneration of bone, cartilage, teeth, and periodontium and treatment of tumors and cysts

ABSTRACT

The present invention provides a composition comprising a corticosteroid and an insulin analog. Further provided is a composition comprising a corticosteroid, insulin lispro and at least one organic acid. The present invention also provides methods of stimulating bone and/or cartilage growth by administering the formulation described herein for the treatment of bone fractures and cartilage damage. Further provided are the methods for the treatment of tumors and cysts of the jaw by administering the formulation described herein.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates generally to the fields of bone andcartilage physiology as well as the treatment of tumors and cysts. Morespecifically, the invention refers to the use of a formulation for thegrowth of bone and cartilage and the treatment of periodontium, tumorsand cysts of the jaw and uses thereof.

2. Description of the Related Art

Osteoarthritis is a degenerative joint disease involving the symptomaticloss of cartilage in load-bearing areas of the joint and is frequentlyfound in the temporomandibular joint, knee, hip, hand, foot and spine.Osteoarthritis causes significant pain and leads to disability and adecrease in the quality of life. Treatment includes orthotics and footwear, physical therapy and exercise, gnathological dental prosthesis andocclusal plates (jaw), pharmacologic therapies such as hyaluronanderivatives and corticosteroids. Surgical management includesosteotomies, partial or total replacement of damaged articulation andautologous chondrocyte implantation, which can only provide ashort-lived reduction in symptoms. Cartilage re-growth may be the idealsolution, but until now, is only used for small defects in specificareas of the femoral cartilage due to the expense and length of timerequired for cartilage re-growth. The cost of surgical treatments isvery high and most of the time, the results do not meet expectations.

The basic management of long bone fractures involves its immobilizationto ensure consolidation. Fracture repair could fail due to severalreasons. For example, pseudarthrosis occurs when bone consolidation isnot achieved after a bone fracture. It is caused by osteotomy orfracture instability, infection and/or loss of bone mass aftercomminuted fractures. Common treatments include physical therapy, pulsedelectromagnetic field, medication with bisphosphonates, curettage ofinvolved bone segments with or without bone graft and circular externalfixation. Osteoporosis is a skeletal disorder characterized bycompromised bone strength that increases the risk of fracture and alsocauses a delay in the consolidation of fractures. Current treatmentincludes the administration of bi- or polyphosphonates, that reduce boneresorption and bone turnover with side effects such as esophageal orgastric irritation, osteonecrosis of the jaw and atypical femoralfractures. Thus, new effective and non-harmful alternative treatmentsare needed.

The goals in treating midface fractures are to restore harmoniousocclusion between the maxilla and mandible to obtain postoperativechewing function and reestablish mid-facial height and facial symmetry.In midface and mandibular fractures, dental occlusion is the mostimportant parameter in reestablishing facial contour and reduction ofthe fracture. Fracture of the alveolar process is a common injury andnearby soft tissues and teeth are often damaged. The aim ofdentoalveolar fracture treatment is to re-establish the normal form andfunction of the masticatory system. Until now, the only treatment forcompromised pulp and/or nerve of the teeth is a root canal, and inrecurrent or more severe cases, dental extraction.

Tumors and cysts of the jaw are very common, have a high rate ofrecurrence, and are classified as odontogenic and non-odontogenic. Amongthe odontogenic cysts, follicular (dentigerous) cysts are particularlyaggressive as they cause root resorption of adjacent teeth in more than50% of cases. A follicular cyst is composed of a membranous sac orcapsule that destroys adjacent bone tissue. Odontoma, a commonodontogenic tumor, frequently interferes with teeth eruption.

Ameloblastoma is a benign but locally aggressive odontogenic tumor withhigh recurrence rates which can occur from the epithelial lining of afollicular cyst. The mandible is commonly affected including theascending ramus, the premolar region and the anterior region, and someameloblastomas are associated with an unerupted tooth. Ameloblastoma isobserved as a radiolucent area presented in a multilocular or unilocularpattern and can be either completely radiolucent or radiolucent withsepta. It causes extensive root resorption and displacement of teeth andhas considerable potential for bone expansion. Surgical treatment mayinvolve resection of the jaw or radiation therapy for inoperable tumors.

Non-odontogenic cysts include aneurysmal bone cysts and globulomaxillarycysts. The aneurysmal bone cyst is a rare benign lesion of thecraniofacial bone which rapidly grows into a bony expansion. Theglobulomaxillary cyst is a fissural cyst which causes divergence ofteeth roots. Treatment consists of surgical curettage or completeexcision of the lesion.

The microenvironment of cysts and tumors has recently become a targetfor new treatments. The tumor's microenvironment is an integral part ofits anatomy, physiology and functionality and is made of stem cells,soluble factors, signaling molecules, and extracellular matrix.Furthermore, the feasibility of differentiating stem cells intospecialized cells or tissues by mechanical stimuli and/or chemical andbiological substances has become evident, especially for tissueengineering applications.

Tissue engineering has the aim of inducing specific cell proliferationor to reprogram cells to heal or achieve tissue regeneration forclinical applications. Tissue engineering techniques include inductivetissue engineering (local delivery of growth factors to stimulate localcells and induce regeneration), conductive tissue engineering (use of amatrix or biomaterials to stimulate growth of existing tissue, celltransplantation (direct introduction of tissue previously manipulated invitro, and hybrid tissue engineering (implantation of an in vitro“prepared” biomaterial within stem cells and growth factors).

Currently, there is no tissue engineering technique, formulation, orminimally invasive methodology or clinical procedure that works with theintralesional microenvironment to ensure even partial regeneration ofinjured bone, cartilage, teeth or periodontium or treatment of tumoralor cystic lesions of the jaw to promote regeneration of damaged tissues.Thus, the prior art is deficient in formulations and methods achievingthese effects. The present invention fulfills this longstanding need anddesire in the art.

SUMMARY OF THE INVENTION

The present invention is directed to a formulation comprising acorticosteroid and an insulin analog.

The present invention is further directed to a non-surgical andminimally invasive method of stimulating regeneration of injured bone ina subject, comprising the step of contacting injured bone with aformulation comprising a corticosteroid and an insulin analog.

The present invention is further directed to a method non-surgical andminimally invasive method of stimulating regeneration of damagedcartilage in a subject, comprising the step of contacting injuredcartilage with a formulation comprising a corticosteroid and insulinanalog.

The present invention is further directed to a non-surgical andminimally invasive method for the treatment of dentoalveolar fracturesand/or regeneration of teeth and/or periodontium in a subject,comprising the step of contacting damaged teeth and/or periodontium witha formulation comprising a corticosteroid and organic acids.

The present invention is further directed to a non-surgical andminimally invasive method for the treatment of tumors and/or cysts ofthe jaw and/or regeneration of the damaged tissue in a subject,comprising the step of contacting tumors and/or cysts with a formulationcomprising a corticosteroid and organic acids.

The present invention is further directed to a formulation comprising acorticosteroid, insulin lispro, and at least one organic acid.

The present invention is further directed to a formulation comprising acorticosteroid and at least one organic acid.

The present invention is further directed to a non-surgical andminimally invasive method for stimulating growth of hair and/or reducinghair loss in a subject in need of such treatment, comprising the stepcontacting scalp or desired tissue with a formulation comprising acorticosteroid and at least one organic acid.

Other and further aspects, features, benefits, and advantages of thepresent invention will be apparent from the following description of thepresently preferred embodiments of the invention given for the purposeof disclosure.

BRIEF DESCRIPTION OF DRAWINGS

So that the matter in which the above-recited features, advantages andobjects of the invention, as well as others that will become clear, areattained and can be understood in detail, more particular descriptionsof the invention briefly summarized above may be had by reference tocertain embodiments thereof that are illustrated in the appendeddrawings. These drawings form a part of the specification. It is to benoted, however, that the appended drawings illustrate preferredembodiments of the invention and therefore are not to be consideredlimiting in their scope.

FIGS. 1A-1B are x-ray images of an atrophic femoral pseudoarthrosislesion that developed over 13 years. FIG. 1A is an x-ray of the femurbefore treatment. FIG. 1B is an x-ray image of the femur fracture shownin FIG. 1A twenty days post-treatment. Osteogenesis in the middle of thefracture can be seen clearly.

FIGS. 2A-2C are x-ray images of a patient with facial fractures. FIG. 2Ashows a panoramic x-ray of jaws with both a parasymphysis and a ramusfracture before treatment. FIG. 2B shows tomography twenty dayspost-treatment revealing consolidation of the pharasymphysis fractureshown in FIG. 2A. FIG. 2C shows tomography twenty days post-treatmentrevealing consolidation of the mandibular ramus fracture shown in FIG.2A.

FIGS. 3A-3B are panoramic x-rays of a patient with exodontia andosteoporosis. FIG. 3A is the inferior first right molar tooth with anacute infection and submaxillary cellulitis by panoramic x-ray.Eosinophilic granuloma is seen in the mesial radicular area of thetooth. FIG. 3B is a panoramic x-ray ten days post-treatment of acuteinfection in the root fragments of the inferior first right molar tooth.Osteogenesis is seen around the extraction area, including low densityregions due to osteoporosis.

FIGS. 4A-4B are panoramic x-rays of a fracture in the ascending ramus ofthe mandible of a patient with post-surgical osteomyelitis of 4 years.FIG. 4A is a panoramic x-ray of the injury after antiseptic disinfectionand before application of the formulation of the present invention andplacement of an external fixator. Notice the severe damage of thecompact bone, the loss of continuity the structure of the bone and thebroken reconstruction plates. FIG. 4B is a panoramic x-ray one monthpost-treatment and placement of an external fixator. Bone growth nearthe right fixator apex (arrow) can be seen clearly.

FIGS. 5A-5C are apical x-rays of a patient with a dentoalveolar fracturewith periodontal tissue damage and an apical cyst. FIG. 5A is an x-rayof a coronal horizontal fracture with major loss of the clinical crownand an apical cyst with bone destruction, loss of periodontal ligamentand rhizolysis before treatment. FIG. 5B is an x-ray seven dayspost-treatment showing osteogenesis. FIG. 5C is an x-ray fourteen daysafter the second treatment with the formulation of the present inventionshowing bone and periodontal ligament regeneration and treatment of theroot resorption. Notice the treatment of the mesial root rhizolysis.

FIGS. 6A-6B are apical x-rays of dental organs with periodontal diseaseand root resorption. FIG. 6A is an x-ray of a patient with periodontaldisease and root resorption before treatment. FIG. 6B is an x-rayfifteen days post-treatment showing regeneration of the damagedperiodontal tissues and injured bone.

FIGS. 7A-7C are x-rays of a patient's left knee with Hallus Valgusarthropathy of 10 years. FIG. 7A is an x-ray of the left knee beforetreatment with the formulation of the present invention. FIGS. 7B-7Cshows an x-ray twenty-one days post-treatment of the left kneearthropahy. Notice the chondrogenesis (arrow) at the site of the injuryin FIG. 7B.

FIGS. 8A-8B are x-rays of a patient with coxarthrosis and post-infectionfemoral bone shortening due to a fracture in the left hip. FIG. 8A is anx-ray of coxarthrosis in the left hip before treatment. FIG. 8B is anx-ray of the left hip one month post-treatment with the formulation ofthe present invention showing chondrogenesis.

FIGS. 9A-9F are x-rays of a dentigerous cyst lesion in the lower rightside of the mandible. FIG. 9A shows tomography of a dentigerous cystbefore treatment. FIG. 9B shows tomography of the cyst in the mandiblebefore treatment. FIGS. 9C-9D show tomography seven days after thesecond treatment with the formulation of the present invention showingan external and internal view of osteogenesis. FIGS. 9E-9F showstomography nine months post-treatment and five months after enucleationof the cyst capsule and extraction of the non-erupted tooth showingosteogenesis of the cortical and cancellous bone.

FIGS. 10A-10F show tomographies of a patient with Gorlin Goltz syndromeand multiple pathologies. FIG. 10A shows tomography of the bilateralmandibular fracture, keratocystic odontogenic tumors, compound andcomplex odontomas and dentigerous cyst. FIGS. 10B-10D show tomography ofconsolidation of both fractures in the mandible six days after treatmentwith the present formulation. Bone growth in the cystic lesions can beseen clearly. FIGS. 10E-10F shows tomography five months post-treatmentwith the present formulation of the bilateral mandibular fracture,keratocystic odontogenic tumors, compound and complex odontomas, anddentigerous cyst. Osteogenesis on the right side of the mandible andfull treatment of the fractures, dentigerous cyst, left keratocysticodontogenic tumor, and complex and compound odontomas.

FIGS. 11A-11D are panoramic radiographies of an odontogenic keratocystictumor and bilateral symphyseal region with multiple calcified areas.FIG. 11A shows radiography of the keratocystic tumor before treatmentwith the formulation of the present invention. FIGS. 11B-11C showradiography seven days post-treatment showing growth of the normalmandibular trabecula along with an increase of bone density. Theosteogenesis of healthy trabecular bone and is denoted by the arrows andcan be seen clearly. FIG. 11D shows tomography of an injured mandiblethree months after treatment. Growth of mandibular trabecula is observedalong with an increase of bone density. Notice the reduction in thehypercalcified regions.

FIGS. 12A-12G are x-rays of a borderline unicystic plexiformameloblastoma in the right hemimandible and a calcifying cystodontogenic tumor (COOT). FIG. 12A is an x-ray before treatment with theformulation of the present invention. Notice the recurrence onpreviously resected bone, including dental organs. FIGS. 12B-12C show anx-ray seven days post-treatment showing neoformation of bony bridges dueto osteogenesis. FIG. 12D is an x-ray fourteen days post-treatmentshowing poor osteogenesis and an extended capsule along the wholelesion, filled with intralesional fluid. Note the appearance of osseoustissue and structures inside the capsule. FIG. 12E shows tomography onemonth after capsule enucleation and the third application of the presentformulation. Notice the filling of the empty trabecular and compact boneniches due to a quick and efficient osteogenesis. FIGS. 12F-12G showtomography 120 days after the third application of the presentformulation showing remarkable osteogenesis.

FIGS. 13A-13B are x-rays of a Gorlin's tumor in the left inferiorpremolar region. FIG. 13A is a panoramic x-ray showing dentoid structurewithin the well formed cystic capsule, before treatment with theformulation of the present invention. FIG. 13B is a periapical x-rayseven days post-treatment showing resorption of the cystic capsule alongwith formation of normal healthy cancellous bone within the injuredarea.

FIGS. 14A-14G show tomography of a multilocular keratocystic odontogenictumor in the right side of the mandible. FIG. 14A is a panoramic x-rayof the injury before treatment with the formulation of the presentinvention. FIG. 14B shows tomography of the right mandibular lingualview of the lesion before treatment with the formulation of the presentinvention. FIG. 14C shows tomography three days post-treatment showingcortical bone regeneration. FIGS. 14D-14E show tomographies three dayspost-treatment showing regeneration of the external cortical andcancellous bone. FIGS. 14F-14G show tomography ninety dayspost-treatment showing treatment of the tumor.

FIGS. 15A-15D show tomographies of a recurrent mandibular aneurysmalbone cyst. FIG. 15A is an x-ray showing bone lysis of the mandibularcondyle and a multilocular lesion in the right ascending ramus. FIG. 15Bis an x-ray before treatment with the formulation of the presentinvention showing a multilocular recurrent lesion and titanium miniplatewith screws from an advanced osteotomy. FIGS. 15C-15D is an x-ray sevendays post-treatment showing almost complete osteogenesis at the site ofthe bone cyst.

FIGS. 16A-16F are x-rays of a central mandibular ossifying fibroma ofeight months. FIG. 16A shows tomography of a fibroma, dental migrationand characteristic bulkiness of the external bone, hyperdense bone areaswith calcifying nodules and low density regions with fluid filledloculi. FIGS. 16B-16C show tomographies of cortical bone bulkinessbefore treatment with the formulation of the present invention. FIG. 16d shows tomography nine days post-treatment showing osteogenesis in theloculi. FIGS. 16E-16F show tomographies thirty five days post-treatmentshowing a decrease of bulkiness of the cortical bone.

FIGS. 17A-17D are x-rays of a globule maxillary cyst in the left side ofthe premaxilla. FIG. 17A is a panoramic x-ray, before treatment with theformulation of the present invention, showing a low density region andextended root displacement which extended to the right side across thebase of the nasal septum. FIG. 17B is an x-ray thirty-five dayspost-treatment showing osteogenesis within the cystic lesion in thepremaxilla. FIGS. 17C-17D is an x-ray sixty-five days post-treatmentshowing osteogenesis of the cancellous and compact bone in the cysticlesion in the premaxilla.

FIGS. 18A-18B are x-rays of a peripheral dentigerous cyst in tooth 13.FIG. 18A is an x-ray before treatment with the formulation of presentinvention showing part of the cystic lesion, labeled with a cross. FIG.18B is an x-ray one month post-treatment showing new bone formation(arrows) within the original cystic lesion in tooth 13.

FIGS. 19A-19E show images of cell cultures. FIG. 19A shows stem cellsisolated from dental pulp. Notice the fibroblastoid morphology. FIG. 19Bshows stem cells isolated from the intralesional content of anameloblastoma. Notice the fibroblastoid morphology. FIG. 19C shows stemcells isolated from an ameloblastoma and treated with one of theformulations of the present invention. Induced proliferation incomparison with the control can be seen. FIG. 19D shows stem cellsisolated from an ameloblastoma and treated with a different formulationof the present invention. A change in the cellular lineage can be seen.FIG. 19E shows control stem cells, isolated from an ameloblastoma andwithout treatment with a formulation of the present invention. Noticethe fibroblastoid morphology.

FIGS. 20A-20B show a 72 year old male with partial alopecia inoccito-parietal area before treatment with Formulation 1 (FIG. 20A) andten days post-treatment (FIG. 20B).

DETAILED DESCRIPTION OF THE INVENTION

As used herein, the following terms and phrases shall have the meaningsset forth below. Unless defined otherwise, all technical and scientificterms used herein have the same meaning as commonly understood to one ofordinary skill in the art.

As used herein, the term, “a” or “an” may mean one or more. As usedherein in the claim(s), when used in conjunction with the word“comprising”, the words “a” or “an” may mean one or more than one. Asused herein “another” or “other” may mean at least a second or more ofthe same or different claim element or components thereof. The terms“comprise” and “comprising” are used in the inclusive, open sense,meaning that additional elements may be included.

As used herein, the term “or” in the claims refers to “and/or” unlessexplicitly indicated to refer to alternatives only or the alternativesare mutually exclusive, although the disclosure supports a definitionthat refers to only alternatives and “and/or”.

As used herein, the term “about” refers to a numeric value, including,for example, whole numbers, fractions, and percentages, whether or notexplicitly indicated. The term “about” generally refers to a range ofnumerical values (e.g., +/−5-10% of the recited value) that one ofordinary skill in the art would consider equivalent to the recited value(e.g., having the same function or result). In some instances, the term“about” may include numerical values that are rounded to the nearestsignificant figure.

As used herein, the term “regeneration” is defined as at least partialosteogenesis and/or chondrogenesis observed, e.g., by imaging and/orx-ray medical techniques as would be readily recognized by a personhaving ordinary skill in this art.

As used herein, the term “teeth” or “tooth” is defined as dentin,cementum and pulp and excludes enamel.

In one embodiment of the present invention, there is provided aformulation, comprising a corticosteroid and an insulin analog.Representative examples of useful corticosteroids include but are notlimited to dexamethasone, cortisol, prednisone, prednisolone,methylprednisolone, betamethasone, triamcinolone, beclometasone,fludrocortisone acetate or deoxycorticosterone acetate. In one preferredembodiment, the corticosteroid is dexamethasone. For example, thedexamethasone may be contained in the formulation in an amount of fromabout 0.5 mg/mL to about 10 mg/mL. A representative example of a usefulinsulin analog is a fast-acting insulin analog. Preferably, thefast-acting insulin analog is insulin lispro. For example, the insulinlispro may be contained in the formulation in an amount of from about0.000115 μg/mL to about 0.00345 μg/mL. Typically, the formulation isdissolved in a vehicle. Representative examples of useful vehiclesinclude but are not limited to normal saline, phosphate buffered salineor water for injection. In one preferred aspect, the formulation furthercomprises at least one organic acid. Representative examples of usefulorganic acids include but are not limited to citric acid, ascorbic acid,malic acid and lactic acid. Generally, such organic acids are containedin the formulation in an amount of from about 0.05 mg/mL to about 5mg/mL. As is well known in the art, the formulation of the presentinvention may be formulated in a variety of forms as pharmaceuticalcompositions or formulations. Representative examples of pharmaceuticalforms include but are not limited to a suspension, spray, solution, gel,paste, ointment, cream, nanoparticle, liposome, microcapsule, deliverydevice or powder.

In another embodiment of the present invention, there is provided anon-surgical and minimally invasive method of stimulating growth orregeneration of bone in a subject, comprising, the steps of contactingthe bone with a formulation comprising a corticosteroid and an insulinanalog, and working with the intralesional microenvironment. Preferably,the growth or regeneration of the bone occurs within a period of time of1 to 6 weeks. Representative examples of useful corticosteroids includebut are not limited to dexamethasone, cortisol, prednisone,prednisolone, methylprednisolone, betamethasone, triamcinolone,beclometasone, fludrocortisone acetate or deoxycorticosterone acetate.In one preferred embodiment, the corticosteroid is dexamethasone. Forexample, the dexamethasone may be contained in the formulation in anamount of from about 0.5 mg/mL to about 10 mg/mL. A representativeexample of a useful insulin analog is a fast-acting insulin analog.Preferably, the fast-acting insulin analog is insulin lispro. Forexample, the insulin lispro may be contained in the formulation in anamount of from about 0.000115 μg/mL to about 0.00345 μg/mL. Typically,the formulation is dissolved in a vehicle. Representative examples ofuseful vehicles include but are not limited to normal saline, phosphatebuffered saline or water for injection. In one preferred aspect, theformulation further comprises at least one organic acid. Representativeexamples of useful organic acids include but are not limited to citricacid, ascorbic acid, malic acid and lactic acid. Generally, such organicacids are contained in the formulation in an amount of from about 0.05mg/mL to about 5 mg/mL. As is well known in the art, this method may beaccomplished using a pharmaceutical composition of the formulation ofthe present invention in a form desirable for this specific method. Aswould be readily recognized by a person having ordinary skill in thisart, this method may be applied to stimulate regeneration of a widevariety of bone or bone-like tissue injures. Representative examples ofbone or bone-like tissue injuries which may be stimulated to regenerateinclude but are not limited to bone fractures, bone fissures, boneresorption, bone necrosis, osteoporosis and pseudoarthrosis.

In yet another embodiment of the present invention, there is provided anon-surgical and minimally invasive method of stimulating theregeneration of damaged cartilage in a subject, comprising the step ofcontacting said injured cartilage with a formulation comprising acorticosteroid and insulin analog, and working with the intralesionalmicroenvironment. Preferably, the regeneration of damaged cartilage isstimulated within a period of time of 1 to 4 weeks. Representativeexamples of useful corticosteroids include but are not limited todexamethasone, cortisol, prednisone, prednisolone, methylprednisolone,betamethasone, triamcinolone, beclometasone, fludrocortisone acetate ordeoxycorticosterone acetate. In one preferred embodiment, thecorticosteroid is dexamethasone. For example, the dexamethasone may becontained in the composition in an amount of from about 0.5 mg/mL toabout 10 mg/mL. A representative example of a useful insulin analog is afast-acting insulin analog. Preferably, the fast-acting insulin analogis insulin lispro. For example, the insulin lispro may be contained inthe formulation in an amount of from about 0.000115 μg/mL to about0.00345 μg/mL. Typically, the composition is dissolved in a vehicle.Representative examples of useful vehicles include but are not limitedto normal saline, phosphate buffered saline or water for injection. Inone preferred aspect, the formulation further comprises at least oneorganic acid. Representative examples of useful organic acids includebut are not limited to citric acid, ascorbic acid, malic acid and lacticacid. Generally, such organic acids are contained in the formulation inan amount of from about 0.05 mg/mL to about 5 mg/mL. As is well known inthe art, this method may be accomplished using a pharmaceuticalcomposition of the formulation of the present invention in a formdesirable for this specific method. As would be readily recognized by aperson having ordinary skill in this art, this method may be applied tostimulate regeneration of a wide variety of cartilage injuries.Representative examples of cartilage injuries include but are notlimited to arthropathies of the temporomandibular joint,acromioclavicular joint, elbow joint, wrist joint, glenohumeral joint,knee joint, coxofemoral joint and meniscal tears.

In yet another embodiment of the present invention, there is provided anon-surgical and minimally invasive method for the treatment ofdentoalveolar fractures and/or regeneration of teeth and/or periodontiumin a subject, comprising the step of contacting the damaged dental organand/or related periodontal tissues with a formulation comprising acorticosteroid and organic acids, and working with the intralesionalmicroenvironment. Preferably, the beneficial treatment of dentoalveolarfractures and/or regeneration of teeth and/or periodontium is seenwithin a period of time of 1 to 2 weeks. Representative examples ofuseful corticosteroids include but are not limited to dexamethasone,cortisol, prednisone, prednisolone, methylprednisolone, betamethasone,triamcinolone, beclometasone, fludrocortisone acetate ordeoxycorticosterone acetate. In one preferred embodiment, thecorticosteroid is dexamethasone. For example, the dexamethasone may becontained in the formulation in an amount of from about 0.5 mg/mL toabout 10 mg/mL. A representative example of a useful insulin analog is afast-acting insulin analog. Preferably, the fast-acting insulin analogis insulin lispro. For example, the insulin lispro may be contained inthe formulation in an amount of from about 0.000115 μg/mL to about0.00345 μg/mL. Typically, the formulation is dissolved in a vehicle.Representative examples of useful vehicles include but are not limitedto normal saline, phosphate buffered saline or water for injection. Inone preferred aspect, the formulation further comprises at least oneorganic acid. Representative examples of useful organic acids includebut are not limited to citric acid, ascorbic acid, malic acid and lacticacid. Generally, such organic acids are contained in the formulation inan amount of from about 0.05 mg/mL to about 5 mg/mL. As is well known inthe art, this method may be accomplished using a pharmaceuticalcomposition of the formulation of the present invention in a formdesirable for this specific method. As would be readily recognized by aperson having ordinary skill in this art, this method may be applied tostimulate the regeneration of a wide variety of injuries in dentalorgans and related periodontal tissues. Representative examples ofinjuries and tissues include but are not limited to, dentoalveolarfractures, root and alveolar bone resorption, rhizolysis, toothmobility, degradation of dentin, cementum, periodontal ligament, pulp,alveolar bone, gingiva and nerve.

In yet another embodiment of the present invention, there is provided anon-surgical and minimally invasive method for the treatment of tumorsand/or cysts of the jaw and regeneration of damaged tissues, comprisingthe step of contacting tumors and/or cysts with a formulation comprisinga corticosteroid and organic acids. Preferably, the beneficial treatmentof tumors and/or cysts of the jaw and regeneration of damaged tissues isseen within a period of 3 to 15 days. Representative examples of usefulcorticosteroids include but are not limited to dexamethasone, cortisol,prednisone, prednisolone, methylprednisolone, betamethasone,triamcinolone, beclometasone, fludrocortisone acetate ordeoxycorticosterone acetate. In one preferred embodiment, thecorticosteroid is dexamethasone. For example, the dexamethasone may becontained in the composition in an amount of from about 0.5 mg/mL toabout 10 mg/mL. Typically, the formulation is dissolved in a vehicle.Representative examples of useful vehicles include but are not limitedto normal saline, phosphate buffered saline or water for injection.Representative examples of useful organic acids include but are notlimited to citric acid, ascorbic acid, malic acid and lactic acid.Generally, such organic acids are contained in the formulation in anamount of from about 0.05 mg/mL to about 5 mg/mL. As is well known inthe art, this method may be accomplished using a pharmaceuticalcomposition of the formulation of the present invention in a formdesirable for this specific method. As would be readily recognized by aperson having ordinary skill in this art, this method may be used totreat a wide variety of tumors and/or cysts of the jaw. Representativeexamples of tumors and/or cysts which may be treated using this methodinclude but are not limited to follicular cysts, keratocysticodontogenic tumors with and without calcifying regions, odontomas,ameloblastomas, Gorlin's tumors, apical cysts, aneurismal bone cysts,ossifying fibromas and globulomaxillary nasolabial cysts. As would bereadily recognized by a person having ordinary skill in this art, thismethod may be used to regenerate a wide variety of damaged tissuescaused by tumors and/or cysts of the jaw. Representative examples oftissues which may be regenerated using this method include but are notlimited to cancellous and compact bone, nerves, gingiva, dentin,cementum, periodontal ligament, pulp, alveolar bone, muscle andcartilage.

In yet another embodiment of the present invention, there is provided anon-surgical and minimally invasive method for stimulating growth ofhair and/or reducing hair loss in a subject in need of such treatment,comprising the step contacting scalp with a formulation comprising acorticosteroid and at least one organic acid. Preferably, the growth ofhair is seen within a period of 7 to 15 days. Representative examples ofuseful corticosteroids include but are not limited to dexamethasone,cortisol, prednisone, prednisolone, methylprednisolone, betamethasone,triamcinolone, beclometasone, fludrocortisone acetate ordeoxycorticosterone acetate. In one preferred embodiment, thecorticosteroid is dexamethasone. For example, the dexamethasone may becontained in the composition in an amount of from about 0.5 mg/mL toabout 10 mg/mL. Typically, the formulation is dissolved in a vehicle.Representative examples of useful vehicles include but are not limitedto normal saline, phosphate buffered saline or water for injection.Representative examples of useful organic acids include but are notlimited to citric acid, ascorbic acid, malic acid and lactic acid.Generally, such organic acids are contained in the formulation in anamount of from about 0.05 mg/mL to about 5 mg/mL. As is well known inthe art, this method may be accomplished using a pharmaceuticalcomposition of the formulation of the present invention in a formdesirable for this specific method. As would be readily recognized by aperson having ordinary skill in this art, this method may be used totreat a wide variety of conditions related with hair loss.Representative examples of conditions related with hair loss which maybe treated using this method include but are not limited to alopecia.

In another embodiment of the present invention, there is provided aformulation, referred to herein as formulation 1, comprising acorticosteroid and at least one organic acid. Representative examples ofuseful corticosteroids include but are not limited to dexamethasone,cortisol, prednisone, prednisolone, methylprednisolone, betamethasone,triamcinolone, beclometasone, fludrocortisone acetate ordeoxycorticosterone acetate. In one preferred embodiment, thecorticosteroid is dexamethasone. For example, the dexamethasone may becontained in the formulation in an amount of from about 0.5 mg/mL toabout 10 mg/mL. Typically, the formulation is dissolved in a vehicle.Representative examples of useful vehicles include but are not limitedto normal saline, phosphate buffered saline or water for injection.Representative examples of useful organic acids include but are notlimited to citric acid, ascorbic acid, malic acid and lactic acid.Generally, such organic acids are contained in the formulation in anamount of from about 0.05 mg/mL to about 5 mg/mL. As is well known inthe art, the formulation of the present invention may be formulated in avariety of forms as pharmaceutical compositions. Representative examplesof pharmaceutical forms include but are not limited to a suspension,spray, solution, gel, paste, ointment, cream, nanoparticle, liposome,microcapsule, delivery device or powder.

In yet another embodiment of the present invention, there is provided aformulation, referred to herein as formulation 2, comprising acorticosteroid; insulin lispro; and at least one organic acid.Representative examples of useful corticosteroids include but are notlimited to dexamethasone, cortisol, prednisone, prednisolone,methylprednisolone, betamethasone, triamcinolone, beclometasone,fludrocortisone acetate or deoxycorticosterone acetate. In one preferredembodiment, the corticosteroid is dexamethasone. For example, thedexamethasone may be contained in the formulation in an amount of fromabout 0.5 mg/mL to about 10 mg/mL. A representative example of a usefulinsulin analog is a fast-acting insulin analog. Preferably, thefast-acting insulin analog is insulin lispro. For example, the insulinlispro may be contained in the formulation in an amount of from about0.000115 μg/mL to about 0.00345 μg/mL. Typically, the formulation isdissolved in a vehicle. Representative examples of useful vehiclesinclude but are not limited to normal saline, phosphate buffered salineor water for injection. In one preferred aspect, the formulation furthercomprises at least one organic acid. Representative examples of usefulorganic acids include but are not limited to citric acid, ascorbic acid,malic acid and lactic acid. Generally, such organic acids are containedin the formulation in an amount of from about 0.05 mg/mL to about 5mg/mL. As is well known in the art, the formulation of the presentinvention may be formulated in a variety of forms as pharmaceuticalcompositions. Representative examples of pharmaceutical forms includebut are not limited to a suspension, spray, solution, gel, paste,ointment, cream, nanoparticle, liposome, microcapsule, delivery deviceor powder.

As is well known in the art, the compositions and formulations of thepresent invention may be applied or administered in a wide variety oftherapeutically effective forms and techniques. In a preferredembodiment, the formulations of the present invention are administeredor applied intralesionally or directly into the injury niche, in orderto work with the content of the microenvironment of the lesion,including stem cells. For example, the formulation of the presentinvention may be applied intralesionally into the periosteal andendosteal injured region for the treatment of bone fractures andosteoporosis. In another embodiment, the formulation of the presentinvention may be applied intralesionally into the niche of the damagedcartilage for arthropathy of the temporomandibular joint, theacromioclavicular joint, the elbow joint, the wrist joint, theglenohumeral joint, the knee or for coxarthrosis. In another embodiment,the formulation of the present invention may be applied intralesionallyand peripherally to compromised teeth and/or compromised periodontaltissues for its treatment and/or regeneration. In yet anotherembodiment, the formulation of the present invention may be appliedintralesionally into the tumors or cysts of the jaw for its treatmentand regeneration of damaged tissues. In another embodiment, theformulation of the present invention may be applied intradermally intothe scalp for the treatment of, for example, alopecia.

As is well known in the art, the compositions and formulations of thepresent invention may be applied or administered to either human ornon-human subjects.

As is well known in the art, the compositions and formulations of thepresent invention may be applied or administered alone or in combinationwith one or more other therapeutic agents, to a subject to treat aparticular condition.

The following examples are given for the purpose of illustrating variousembodiments of the invention and are not meant to limit the presentinvention in any fashion.

Example 1 Treatment Method for Cysts and Tumors of the Jaw

When present, most of the intralesional fluid is drained. If clinicalevidence of infection is found inside the lesion, the niche is washed.Depending on the size of the injury, from 1-8 mL of an antiseptic(electrolyzed superoxidized water solution of neutral pH, ESTERICIDE®)is injected and immediately drained back. Formulation 1 of the presentinvention is applied, intralesionally and percutaneously, perfusing allloculi, making sure to reach all perimeter walls of the lesion,according to the tomography, and, if necessary, a fluoroscope is used.Depending on the tumor's size, 0.5-5.0 mL of the formulation may beperfused. The progress of the treatment is clinically monitored every 3or 5 days, any intralesional fluid is aspirated with a needle, andtomographic follow-up is given one and two weeks post-treatment.

Three cases may arise. First, if the osteogenesis progressedsatisfactorily, clinical follow-up is continued weekly with a monthlytomography (during the first two months and tomography is performedevery three months), until observing treatment of the lesion. Second, ifthe osteogenesis stops completely or slowly progresses, with or withoutformation of intralesional fluid, then the intralesional fluid isaspirated with a needle, if formed, and a second application of thepresent formulation is performed. The patient is monitored every 3 or 5days along with aspiration of any intralesional liquid with a needle, ifformed, and a monthly tomography (during the first two months and thenthe tomography is performed every three months) until the lesion hasbeen completely resolved. Third, if bone resorption occurs with orwithout formation of a cystic or tumoral capsule with or withoutintralesional liquid, then enucleation of the capsule is performedand/or curettage of the injury under local anesthesia, the wound isclosed and the formulation of the present invention is applied. Thepatient is monitored every 3 or 5 days with aspiration of anyintralesional liquid, if formed, and monthly tomography is performed(during the first two months and then the tomography is performed everythree months) until the lesion has completely resolved. In all cases,after observing treatment of the injury, clinical monitoring iscontinued in accordance with the regulations for this type of pathology.

Example 2 Treatment Method for Regeneration of Teeth and Periodontium

Dental cleaning (scaling) is performed using ESTERICIDE® antiseptic asthe irrigant. In the case of periodontal disease or infection, anantiseptic (ESTERICIDE®) is perfused intralesionally and peripheral tothe compromised tooth, once a day until eradication of the infection. Incase of tooth decay, cavity, dental crack, loose filling or pulpexposure, the injury is treated first. When major loss of dental tissuehas occurred, significant mobility may be present, so ferulization mightbe required. Peripheral immobilization with surgical wire could benecessary when treating a coronal and/or apical fractured tooth.Application of sodium fluoride is recommended when treating a fractureinvolving presence of enamel. Approximately 0.5-3.0 mL of formulation 1of the present invention was applied peri- and intraligamentary.

Example 3 Treatment Method for Bone Fractures and Osteoporosis

If required, the injury is pretreated as follows. In the case ofosteomyelitis, 1-5 mL of antiseptic (ESTERICIDE®) is perfusedintralesionally, every three days for six weeks. Formulation 2 of thepresent invention is applied after three days. For facial fractureswithout infection, 0.5-4 mL of antiseptic (ESTERICIDE®) is perfusedintralesionally in the fracture bed. For mandibular or long bonefractures, an external fixator is placed adjusting to the correctposition of the bone segments. In all cases, 0.5-5.0 mL of theformulation of the present invention is applied intralesionally into theperiosteal and endosteal region, perfusing into cancellous bone.

Example 4 Treatment Method for Regeneration of Cartilage for Arthropathyof Temporomandibular Joint

Local nerve block anesthesia is performed with antisepsis. To identifythe puncture site, an imaginary line of 2 cm is drawn from the ear'stragus to the eye's lateral canthus and then a perpendicular line of 1cm is drawn from that point to the angle of mandible. After ensuring thecorrect puncture location, 2 mL of formulation 2 of the presentinvention was perfused. A protective dressing of ESTERICIDE® gel isapplied over puncture site. A second application of the presentformulation is administered one month post-treatment with radiographicand tomographic monitoring of the lesion.

Example 5 Treatment Method for Regeneration of Cartilage for Arthropathyof Acromioclavicular Joint

The patient's hands are placed to rest over her/his lap remaining stillwithout moving arms or shoulders, and looking to the opposite directionof the injured joint. Antisepsis and intramuscular infiltration oflidocaine (2%) is performed. The skin entry site is just above theacromioclavicular joint. To locate it, the clavicular line is followedfrom the medial to lateral position and the joint is in a smalldepression zone that presents sensitivity in the case of an injury.After ensuring the correct puncture location, 2 mL of formulation 2 ofthe present invention is perfused. A protective dressing of ESTERICIDE®gel was applied over the puncture site. A second application of thepresent formulation was performed as described within one monthpost-treatment with radiographic and tomographic monitoring of thelesion.

Example 6 Treatment Method for Regeneration of Cartilage for Arthropathyof Elbow Joint

The elbow remains slightly flexed and supported over a pillow with thewrist in a neutral position. With antisepsis, local anesthesia wascutaneously administrated. The skin entry site was in a depression zonelocated immediately proximal to radial head. After ensuring the correctpuncture location, 2 mL of formulation 2 of the present invention isperfused. A protective dressing of ESTERICIDE® gel was applied over thepuncture site. A second application of the formulation of the presentinvention was administered as described before, one monthpost-treatment, with radiographic and tomographic monitoring of thelesion.

Example 7 Treatment Method for Regeneration of Cartilage for Arthropathyof Wrist Joint

The elbow remains slightly flexed while the wrist in rolled inward andin a neutral position. With antisepsis, local anesthesia is cutaneouslyadministrated. The skin entry site is over the dorsal position of thewrist's articulation, just above the site with the highest sensitivity.After ensuring the correct puncture location, 2 mL of formulation 2 ofthe present invention is perfused. A protective dressing of ESTERICIDE®gel is applied over the puncture site. A second application of theformulation of the present invention is administered as describedbefore, one month post-treatment, with radiographic and tomographicmonitoring of the lesion.

Example 8 Treatment Method for Regeneration of Cartilage for Arthropathyof Glenohumeral Joint

The procedure must be performed with an anterior approach. The patient'shands are placed to rest over his/her lap remaining still without movingthe arms or shoulders. With antisepsis, intramuscular infiltration oflidocaine (2%) is performed. The skin entry site is just below andmedial to the posterolateral aspect of the acromion, and from thisposterior approach, the needle tip is directed towards the coracoidprocess. After ensuring the correct puncture location, 2 mL offormulation 2 of the present invention was perfused. A protectivedressing of ESTERICIDE® gel was applied over the puncture site. A secondapplication of the present formulation was administered as describedbefore, one month post-treatment, with radiographic and tomographicmonitoring of the lesion.

Example 9 Treatment Method for Regeneration of Cartilage for Arthropathyof Knee

With antisepsis, intramuscular infiltration of lidocaine (2%) wasperformed to the knee in a semiflexed position (45°). The medial borderof the patellar tendon was located and a puncture with an hypodermicneedle was performed, perpendicularly to skin, immediately medial topatella until reaching the articular and synovial capsule. Synovialfluid was extracted to assure the correct location of the puncture and 2mL of formulation 2 of the present invention was perfused. A protectivedressing of ESTERICIDE® gel was applied over the puncture site. A secondapplication of the present formulation was administered as describedabove, one month post-treatment, with radiographic and tomographicmonitoring of the lesion.

Example 10 Treatment Method for Regeneration of Cartilage forCoxarthrosis

Sedation of the patient was performed as well as antisepsis of thesurgical area. The puncture site is located in the coxofemoral jointregion, the major vortex of trochanter was situated approximately 6 cmstraightforward to imaginary middle line of body and 6 cm from there toa cephalic direction. With the use of fluoroscopy, the needle insertionsite was located and a blocking needle is introduced to perfuse 2 mL offormulation 2 of the present invention. A protective dressing ofESTERICIDE® gel was applied over the puncture site. A second applicationof the present formulation was administered as described before, onemonth post-treatment, with radiographic and tomographic monitoring ofthe lesion.

Example 11 Treatment of Long Bone (Femur) Fracture

A 26 year old male patient with 12 previous surgeries presented with 13year-old atrophic femoral pseudoarthrosis. His x-ray revealed a femurfracture supported by a fixation plate and ten screws (FIG. 1A). Underpercutaneous local anesthesia and antisepsis with ESTERICIDE®antiseptic, 2 mL of the present formulation 2 were perfusedintralesionally using a fluoroscope and peripheral nerve block catheter.Finally, a protective dressing of antiseptic ESTERICIDE® gel was placedon the puncture site. Twenty days post-treatment, a second applicationof the present formulation was administered. Forty days after the firsttreatment, radiographic imaging showed clear osteogenesis in the centerof fracture (FIG. 1B).

Example 12 Treatment of Double Facial (Parasymphysis and MandibularRamus) Fracture

A 25 year old male patient presented with facial fractures. His x-rayrevealed both a parasymphysis fracture and a fracture of the leftmandibular ramus (FIG. 2A). Under general anesthesia and antisepsis withESTERICIDE®, external fixators were placed and 4 mL of the presentformulation 2 (2 mL in each fracture niche) were perfusedintralesionally using a fluoroscope. Finally, a protective dressing ofESTERICIDE® gel was placed on the puncture site. Twenty dayspost-treatment, the external fixators were removed and radiographicimaging showed consolidation of both fractures (FIGS. 2B-2C).

Example 13 Treatment of Tooth Extraction and Osteoporosis

A 61 year old female patient with osteoporosis, controlled hypertensionand diabetes presented with an acute infection in the root fragments ofthe inferior first right molar tooth with submaxillary cellulitis (FIG.3A). The patient insisted her tooth be removed but accepted treatmentwith the formulation of the present invention to repair the bone defectdue to extraction and osteoporosis. After eradication of the infection,extraction of the tooth was performed under local anesthesia and thesurgical bed was washed with antiseptic. Subsequently, an aspirationneedle filled with 2 mL of formulation 2 was introduced to the surgicalbed until cancellous bone was reached and the formulation was slowlyperfused.

Osteogenesis was observed by radiographic imaging, ten dayspost-treatment (FIG. 3B). An important increment of bone density wasobserved not only in the extraction zone but within the zone ofosteoporosis damage so she was referred after one month for clinical andradiographic monitoring. Thus, it was demonstrated that the formulationof the present invention is a therapeutically effective treatment forosteoporosis disease.

Example 14 Treatment of Osteomyelitis, Osteoporosis and Facial Fracture

A 60 year old female patient presented with osteoporosis, post-surgicalosteomyelitis of 4 years and a fracture in the ascending ramus of themandible. The patient also had a neoplasia in the current injured zoneand osteomyelitis occurrence after its enucleation and curettage.

3 mL of the antiseptic solution ESTERICIDE® was applied intralesionally,two times per week, for six weeks and oral antibiotics were alsoprescribed which eradicated the infection. An initial panoramic x-rayshowed severe bone degradation (FIG. 4A). After eradication ofosteomyelitis and under general anesthesia, 3 mL of the presentformulation 2 were perfused intralesionally, using fluoroscopy. Then, anexternal fixator was placed for stabilization of the mandible. Theexternal fixator is a substitute for the reconstruction plates whichmust be placed with the surgical procedure. Finally, a protectivedressing of superoxide electrolyzed solution and gel was placed over theexternal wounds.

Fifteen days post-treatment, clinical and radiographic monitoringrevealed an increase in bone density around the injury. One monthpost-treatment, (FIG. 4B) a panoramic image revealed the formation ofnew bone close to the right fixator apex (arrow) with no recurrence ofosteomyelitis. A second application of the present formulation was madeas described above. After one month, the patient was monitoredclinically and radiographically which showed that the procedureeliminated 4 year old osteomyelitis and induced osteogenesis.

Example 15 Treatment of Dentoalveolar Fracture with Periodontium Damageand Apical Cyst with Bone Destruction

A 28 year old male patient with a root canal of tooth 30, presented witha coronal horizontal fracture, major loss of the clinical crown and anapical cyst with bone destruction, loss of the periodontal ligament andrhizolysis (FIG. 5A). The dental organ was severely damaged andexodontia was necessary but the patient agreed to treatment withformulation 1 of the present invention.

The treatment was performed under local mandibular nerve blockanesthesia (Mevipacaine; 1 mL; 3%), antisepsis with ESTERICIDE®. Then,an aspiration needle was used to extract less than 0.5 mL of bloodyexudate by negative aspiration and 2 mL of the present formulation wereslowly perfused. Seven days post-treatment, radiography showedosteogenesis and regeneration of the horizontal coronal fracture (FIG.5B). A second dosage of the present formulation was administrated andradiography fourteen days post-treatment showed bone regeneration,treatment of root resorption as well as regeneration of the periodontalligament, treatment of the mesial root rhizolysis gaps (FIG. 5C).

Example 16 Treatment of Dental Organs with Periodontal Disease and RootResorption

A 78 year old male patient presented with periodontal disease in theanterior dental organs with superior and middle third root resorption oftooth 44 and tooth 42 (FIG. 6A). The dental organs were severely damagedand exodontia was mandatory. The patient agreed to treat his teeth withformulation 1 of the invention.

Under local mandibular nerve block anesthesia, scaling was performedalong with antisepsis with ESTERICIDE®. Then, 2 mL of the formulation ofthe present invention were applied peri- and intraligamentarily. Fifteendays post-treatment, there was full regeneration of the damaged teethand osteogenesis of the injured bone (FIG. 6B). Thus, the formulation ofthe present invention was able to regenerate the damaged bone andperiodontal ligament and also dentin and cement.

Example 17 Treatment of Left Knee Arthropathy with CartilageRegeneration

A 84 year old female presented with Hallus Valgus left knee arthropathyof 10 years (FIG. 7A). After antisepsis of the knee surface, lidocaine(2%) was administered via intramuscular infiltration of the knee in asemiflexed position (45°). Then, 2 mL of the present formulation wereinjected in the medial border of the patellar tendon perpendicular tothe skin, immediately medial to the patella reaching the articular andsynovial capsule. Formulation 2 of the present invention was slowlyperfused and a protective dressing of ESTERICIDE® gel was applied overthe puncture site. After 21 days, the patient showed remarkableimprovement in walking and knee flexibility, as well as chondrogenesis(FIGS. 7B-7C). A second application of the formulation was administereda week later as described. After 4 months, the patient's mobility andflexibility was significantly improved.

Example 18 Treatment of Coxarthritis with Cartilage Regeneration

A male patient presented with coxarthrosis and post-infection femoralbone shortening due to a fracture in the left hip (FIG. 8A). Therecommended treatment is arthroplasty and vigorous physicalrehabilitation. The patient, however, agreed to treat his pathology withformulation 2 of the present invention. After sedation and antisepsis, 2mL of the present formulation was administered into the coxofemoraljoint region via fluoroscopy. The formulation was slowly perfused and aprotective dressing of ESTERICIDE® gel was applied over the puncturesite. One month post-treatment, there was a remarkable improvement ofmovement range and chondrogenesis at the site of the injury (FIG. 8B).

Example 19 Treatment of Dentigerous Cyst

A 21 year old male patient presented with slight hyperaesthesia and aright mandibular osteolytic lesion. Radiolography showed a multilocularlesion with rhizolysis, a defined capsule and destruction of theexternal and internal cortical bone, encompassing the body and ascendingramus of the mandible. There was a developing dental organ in theectopic position, near the mandibular basal bone with a clinical crownand rhizolysis in the apical third of the second lower right molar(FIGS. 9A-9B). Typical treatments include marsupialization, subsequentremoval of the cystic capsule, enucleation and curettage of theneoplasia, which has a risk of recurrence. The patient agreed to treathis pathology with formulation 1 of the present invention.

Local anesthesia (Mepivacaine; 1 mL; 3%) was given subdermally in theright pre-auricular region after antisepsis with ESTERICIDE®.Subsequently, an aspiration needle was used to extract 9 mL of theintracystic liquid. Then, 2 ml of the formulation of the presentinvention was slowly perfused. A protective dressing of antisepticelectrolyzed solution and gel was placed over the puncture site.

Seven days post-treatment, radiography revealed intra- andextra-lesional osteogenesis and resorption of the bone was observedafter one month due to the presence of the non-erupted dental organ andthe multilocular nature of the lesion. Therefore, a second applicationof the present formulation was performed two months after the firstapplication. Tomography seven days later showed regeneration of theexternal cortical bone and the cancellous bone (FIGS. 9C-9D). Two monthspost-treatment, extraction of the non-erupted tooth along with itscystic capsule was performed and after five months, tomography showedregeneration and remodeling of the injury (FIGS. 9E-9F).

Example 20 Treatment of Bilateral Mandibular Fracture, OdontogenicTumors and Cyst

A 19 year old female with perinatal cerebral hypoxia and Gorlin Goltzsyndrome presented with a bilateral mandibular fracture, twokeratocystic odontogenic tumors, three odontomas (compound, complex andcystic) and a dentigerous cyst (FIG. 10A). Placement of reconstructionplate and ulterior iliac bone graft along with curettage or enucleationof remaining cystic lesions was highly recommended. Informed consent(Helsinki's protocol) for formulation 1 of the present invention wasobtained.

Under general anesthesia, after antisepsis with ESTERICIDE®, anaspiration needle with a 20 mL syringe was introduced intralesionally inthe right keratocystic tumor, 4 mL of seropurulent exudate was extractedwith negative pressure and the lesion was washed with an antisepticsolution. Finally, 2 mL of the present formulation were perfused intothe lesion. Next, an aspiration needle with a 10 mL syringe wasintroduced intralesionally in the dentigerous cyst, 2 mL of a denseyellowish fluid was extracted by suction, and 1 mL of formulation 1 wasslowly perfused into the cyst.

For treatment of the odontomas, 1 mL of serohematic fluid was extractedfrom each mandibular odontoma and 0.5 mL of the present formulation wasslowly perfused into each odontoma. The complex odontoma, located on theleft side of the maxilla, was treated with 1 mL of the presentformulation after extraction of 0.5 mL of serohematic fluid.

Finally, a protective dressing of an antiseptic electrolyzed solutionand gel was placed over the puncture site. Five days post-treatment,radiography revealed full consolidation of the fractures and clearevidence of a decrease of the cystic lesions (FIGS. 10B-10D). Thefixator was removed and enucleation of the right side of thekeratocystic capsule was performed. Finally, 1 mL of the presentformulation was perfused into the lesion and a protective dressing ofsuperoxide gel was applied. The patient was monitored five monthspost-treatment (FIGS. 10E-10F) and had osteogenesis in the right side ofthe mandible and treatment of fractures, dentigerous cyst, keratocysticodontogenic tumor and odontomas.

Example 21 Treatment of Keratocystic Odontogenic Tumor with CalcifyingAreas

A 22 year old male presented with pain in the right retromolar space andpericoronitis at the third molar. Laboratory tests were normal butpanoramic radiography showed diffuse mandibular images with radiolucentand radiopaque regions in the body and the symphyseal area (FIG. 11A).Histopathological examination revealed a keratocystic odontogenic tumorcongruent with nevoid basal cell carcinoma (Gorlin) syndrome. Thepatient agreed to treatment with formulation 1 of the present invention.

After antisepsis with ESTERICIDE®, local mandibular nerve blockanesthesia (1.8 mL of Dentocaine®: Mepivacaine 36 mg, epinephrine 18 mg)was applied. The lesion was punctured with an aspiration needle betweenthe inferior second premolar and first molar region. The extraoralapproach was made under the bottom of the vestibular sac, avoiding anycontact with the oral cavity and making rotational movements with theneedle until penetration of the vestibular compact bone. After thebiopsy, the present formulation was introduced intralesionally andslowly perfused. Finally, a protective dressing of an antisepticelectrolyzed solution and gel was placed over the puncture.

Tomography seven days post-treatment (FIGS. 11B-11C) showed regenerationof mandibular trabecula without the appearance of hypercalcified areasand an increase in bone density. Three months post-treatment, tomography(FIG. 11D) showed osteogenesis of healthy cancellous bone at the injury.

Example 22 Treatment of Borderline Ameloblastoma with Calcifying CystOdontogenic Tumor (COOT)

A 26 year old patient presented with paresthesia, a borderline unicysticplexiform ameloblastoma of the mandible and a calcifying cystodontogenic tumor in the right hemimandible (FIG. 12A). The patientagreed to treatment with formulation 1 of the present invention.

Treatment was made under bilateral nerve block anesthesia withMepivacaine 3%, after antisepsis with ESTERICIDE®. A needle with syringewas inserted intralesionally, via a mental and submental percutaneousapproach, for the aspiration of 37 mL of serohematic exudate. With thesame procedure, 4 mL of the present formulation were injected: 2 mL inthe back side of tumor and 2 mL in the mental area of the tumorperfusing into all of the intralesional locules so as to reach theperimeter of the lesion.

Three days later, 20 mL of intralesional exudate were extracted asdescribed above. Seven days post-treatment, tomography (FIGS. 12B-12C)showed remarkable bone regeneration, including bony bridges and externalcortical bone.

Three days later, 3 mL of intralesional exudate were extracted and asecond application of the present formulation was performed as describedabove. Radiography five days later showed no metastasis but littleresorption of the neoformed bone and no further osteogenesis wasobserved. In contrast, a tumor capsule within dentoid structures andbone spicules was observed to be extended over the lesion (FIG. 12D).The capsule was removed after one month and a third intralesionalapplication of the present formulation (2 mL) was made. One month later,tomographies showed osteogenesis at the lesion (FIG. 12E). Three monthsafter the third application of the present formulation, a remarkableosteogenesis (95%) was observed (FIGS. 12F-12G).

Example 23 Treatment of Gorlin's Tumor

A 17 year old male presented with a cystic lesion comprised of adentoid-like calcification within a well formed cystic capsule, locatedin the apical and half region of the left inferior premolars, congruentwith a Gorlin's tumor (FIG. 13A). Treatment includes enucleation of thecystic capsule but the patient agreed to treatment with formulation 1 ofthe present invention.

After antisepsis with ESTERICIDE®, local mandibular nerve blockanesthesia (Mepivacaine; 1 mL; 3%) was administered. Then, 1 mL of theformulation of the present invention was intralesionally slowlyperfused. Finally a protector deposit of an antiseptic electrolyzedsolution and gel was placed over the puncture site. Seven dayspost-treatment, a periapical radiography showed complete boneregeneration of the injured zone with absence of a peripheral cysticcontour (resorption of capsule) along with normal trabecular boneformation (FIG. 13B).

Example 24 Treatment of Keratocystic Odontogenic Tumor

A 20 year old male presented with a mandibular cyst. Radiography showeda lytic mandibular multilocular lesion on the right side withintralesional liquid, comprising the ascending ramus, body, condyle,coronoid apophysis and premolars (FIGS. 14A-14B). A keratocysticodontogenic tumor after biopsy, and the patient agreed to treatment withformulation 1 of the present invention.

The patient had mandibular nerve block anesthesia (Mepivacaine; 1 mL;3%) and antisepsis with ESTERICIDE®. Subsequently, an aspiration needlewas used to extract by negative aspiration 25 mL of brown waxyintracystic content. Then, 2 mL of the formulation of the presentinvention was slowly perfused intralesionally. Finally, a protectivedressing of electrolized gel was placed over the puncture site closed bya stitch. Three days post-treatment, tomography showed full regenerationof cortical and cancellous bone (FIGS. 14C-14E).

Eighty days post-treatment, the inner face of the condile showed partialregeneration, so 8 mL of a dark brown exudate were extracted and asecond application of the present formulation was performed. Seven dayspost-treatment, tomography showed complete eradication of the cysticlesion and osteogenesis with regeneration of the injured tissues (FIGS.14F-14G).

Example 25 Treatment of Recurrent Aneurysmal Bone Cyst

A 13 year old male presented with a recurrent aneurysmal bone cyst ofthe mandible. The patient had two prior surgeries so an osteotomy wasperformed to cover the previous bone defect. Initial tomography showed alytic lesion of the condyloid process of the mandible and a multilocularcystic region (with no well defined margins) of osseal hypodensity inthe right ascending ramus (FIG. 15A). His second multiplanar tomographywith reconstruction (FIG. 15B) showed a titanium miniplate (2.0 mm) withscrews to hold the cortical osteotomy and a multilocular recurrentlesion. Typical treatment includes block resection of the rightascending ramus and further reconstruction with revascularizedautologous bone from the iliac crest. The patient's parents agreed totreatment with formulation 1 of the present invention.

Due to the patient's age, the procedure was done under generalanesthesia after antisepsis with ESTERICIDE®. An aspiration needle wasinserted percutaneously and intralesionally into the temporal fossa foraspiration of 10 mL of hematic exudate. Next, 2 mL of formulation 1 ofthe present invention was injected via intralesional puncture. Finally,a protective dressing of an antiseptic electrolyzed solution and gel wasplaced over the puncture spots. Six days post-treatment, the patient'sradiographic monitoring showed that the external cortical bone of thecystic lesion had completely healed and the presence of bone growth(FIGS. 15C-15D).

Example 26 Treatment of Mandibular Ossifying Fibroma

A 35 year old female presented with paresthesia due to a centralmandibular ossifying fibroma of eight months. The lesion was anaugmentation of the cortical bone without damage to the oral mucosa.Radiography revealed dental migration and characteristic bulkiness ofthe external bone tables, hyperdense bone areas with calcifying nodulesand hypodense regions with loculi apparently filled with liquid (FIGS.16A-16C). Recommended treatment is a block resection surgery but thepatient agreed to treatment with formulation 1 of the present invention.

After antisepsis with ESTERICIDE®, local bilateral mandibular nerveblock anesthesia (Mepivacaine; 1 mL; 3%) was administered. Subsequently,an aspiration needle was used to extract 2 mL of serohematic liquid bynegative aspiration. Then, 3 mL of the formulation of the presentinvention was slowly perfused intralesionally. Finally, a protectivedressing of antiseptic electrolyzed solution and gel was placed over thepuncture site. Three days post-treatment a second application offormulation 1 was performed. Six days post-treatment, the paresthesiadisappeared and tomography showed osteogenesis in the loculi (FIG. 16D).Thirty-five days post-treatment, tomography showed diminishment of thelesion's bulkiness (FIGS. 16E-16F).

Example 27 Treatment of Globulomaxillary Cyst

A 25 year old female presented with a globulomaxillary cyst and a lowdensity lesion in the left side of the premaxilla (FIG. 17A).Conventional treatment for this kind of lesion is surgical enucleationwhich has a risk of recurrence. After obtaining informed consent(Helsinki's protocol), formulation 1 of the present invention wasadministered.

After antisepsis with ESTERICIDE®, infraorbital nerve block (1.8 mL ofDentocaine®: Mepivacaine 36 mg, epinephrine 18 mg) and palatal-anterioranesthesia (1.8 mL of Dentocaine® simple) was administered.Subsequently, the lesion was percutaneously punctured across the upperlip and 6 mL of yellowish brown liquid were extracted by negativeaspiration. Then, 2 mL of the formulation of the present invention wasslowly perfused intralesionally. A protective dressing of ESTERICIDE®gel was applied to the puncture sites. Seven days post-treatment, thepatient's tomography showed initial osteogenesis within the cysticlesion. Radiography 14 days post-treatment revealed disappearance of thelesion and improved bone growth over the cystic lesion (FIG. 17B). 65days post-treatment, there was complete osteogenesis within the originalcystic lesion (FIGS. 17C-17D).

Example 28 Treatment of Peripheral Dentigerous Cyst in Tooth 13

A 9 year old male presented with a peripheral dentigerous cyst in tooth13 and radiography showed a cystic lesion (FIG. 18A). The patient agreedto treatment with formulation 1 of the present invention.

Under sedation and local anesthesia and antisepsis with ESTERICIDE®, anaspiration needle was inserted intralesionally and 3 mL of a denseyellowish liquid were extracted by negative aspiration. Then, 2 mL ofthe formulation of the present invention were slowly perfusedintralesionally. Finally, a protective dressing of ESTERICIDE® gel wasapplied at the puncture site. One month post-treatment, radiographyshowed osteogenesis within the original cystic lesion (FIG. 18B).

Example 29 Detection of Stem Cells in the Intralesional Fluid of TreatedBorderline Ameloblastoma and Comparison with Stem Cell Cultures Obtainedfrom Dental Pulp

Stem cells were obtained from dental pulp under conventionalmethodologies and cultured in petri dishes (35 mm×10 mm) in DMEM culturemedium (FIG. 19A). Stem cells from the ameloblastoma were purified byfiltration and centrifugation of the intralesional fluid and culturedunder the above conditions. The obtained cells showed typical stem cellsmorphology (FIG. 19B).

An equal number of the purified stem cells (1 mL) was cultured, aspreviously described, in three separated petri dishes and 1 mL of DMEMmedium was added. Then, 5 μL/mL of formulation 1 of the presentinvention were added to the first dish, 5 μL/mL of formulation 2 of thepresent invention were added to the second dish and the third dishreceived no treatment. After two weeks, there was significantproliferation of the cells in dish one (FIG. 19C), apparentdifferentiation of cells in dish two into chondrocytes (FIG. 19D), and amodest quantity of cells with the typical fibroblastoid morphology inthe control dish (FIG. 19E). Thus, this experiment demonstrated that theformulations of the present invention induce significant proliferationof cells in a short period of time and induce a change in the cellularlineage.

Example 30 Treatment for the Stimulation of Hair Growth

A 72 year old male presented partial alopecia in occito-parietal area(FIG. 20A). The patient agreed to treatment with formulation 1 of thepresent invention. Under local anesthesia and antisepsis withESTERICIDE®, an aspiration needle was inserted intradermically intodifferent areas of the scalp and 1.5 mL of Formulation 1 were perfusedfrom inside to outside, in order to stimulate both cellular niches andcovering 15% of the alopecia area. Finally a protective dressing ofESTERICIDE® gel was applied at the puncture sites. Ten dayspost-treatment, the growth of hair was observed (FIG. 20B). A secondapplication of the present formulation was administered 20 dayspost-treatment in remanent areas of alopecia. Thus, the formulation ofthe present invention may be to stimulate growth of hair and treat hairloss. In addition, as illustrated in FIG. 20B, the color of thepatient's hair darkened after treatment with the formulation of thepresent invention and thus, this formulation may be used to stimulatepigmentation of the hair follicle.

“Stimulating hair growth” includes stimulating an increase in total hairmass and/or length. Such increase includes increased length and/orgrowth rate of hair shafts (i.e. follicles), increased number of hairs,and/or increased hair thickness. “Stimulating hair growth” should alsobe considered to include preventing, arresting, decreasing, delayingand/or reversing hair loss. As is readily apparent, the presentinvention is further directed to a method for stimulating growth of hairand/or reducing hair loss in a subject in need of such treatment,comprising the step of contacting scalp with a formulation comprising acorticosteroid and at least one organic acid.

The present invention is well adapted to attain the ends and advantagesmentioned as well as those that are inherent therein. The particularembodiments disclosed above are illustrative only, as the presentinvention may be modified and practiced in different but equivalentmanners apparent to those skilled in the art having the benefit of theteachings herein. Furthermore, no limitations are intended to thedetails of construction or design herein shown, other than as describedin the claims below. It is therefore evident that the particularillustrative embodiments disclosed above may be altered or modified andall such variations are considered within the scope and spirit of thepresent invention.

What is claimed is:
 1. A formulation, comprising: a corticosteroid, andinsulin lispro in an amount of about 0.000115 μg/mL to about 0.00345μg/mL.
 2. The formulation of claim 1, wherein said corticosteroid isdexamethasone, cortisol, prednisone, prednisolone, methylprednisolone,betamethasone, triamcinolone, beclometasone, fludrocortisone acetate ordeoxycorticosterone acetate.
 3. The formulation of claim 2, wherein saiddexamethasone is contained in said formulation in an amount of fromabout 0.5 mg/mL to about 10 mg/mL.
 4. The formulation of claim 1,wherein said formulation is dissolved in a vehicle.
 5. The formulationof claim 1, further comprising at least one organic acid selected fromthe group consisting of citric acid, ascorbic acid, malic acid andlactic acid.
 6. The formulation of claim 5, wherein said at least oneorganic acid is contained in said formulation in an amount of from about0.05 mg/mL to about 5 mg/mL.
 7. The formulation of claim 6, wherein saidformulation is a pharmaceutical composition.
 8. A method of stimulatingbone growth in a subject, comprising the steps of: contacting said bonewith a formulation comprising a corticosteroid and insulin lispro in anamount of about 0.000115 μg/mL to about 0.00345 μg/mL.
 9. The method ofclaim 8, wherein said corticosteroid is dexamethasone, cortisol,prednisone, prednisolone, methylprednisolone, betamethasone,triamcinolone, beclometasone, fludrocortisone acetate ordeoxycorticosterone acetate.
 10. The method of claim 9, wherein saiddexamethasone is contained in said formulation in a concentration rangeof from about 0.5 mg/mL to about 10 mg/mL.
 11. The method of claim 8,wherein said formulation contains at least one organic acid.
 12. Themethod of claim 11, wherein said at least one organic acid is citricacid, ascorbic acid, malic acid or lactic acid.
 13. The method of claim12, wherein said at least one organic acid is contained in saidformulation in a concentration range of about 0.05 mg/mL to about 5mg/mL.
 14. A method of stimulating cartilage growth in a subject,comprising the step of: contacting said cartilage with a formulationcomprising a corticosteroid and insulin lispro in an amount of about0.000115 μg/mL to about 0.00345 μg/mL.
 15. The method of claim 14,wherein said corticosteroid is selected from the group consisting ofdexamethasone, cortisol, prednisone, prednisolone, methylprednisolone,betamethasone, triamcinolone, beclometasone, fludrocortisone acetate,deoxycorticosterone acetate and aldosterone.
 16. The method of claim 14,wherein said formulation contains at least one organic acid.
 17. Themethod of claim 16, wherein said at least one organic acid is citricacid, ascorbic acid, malic acid or lactic acid.